Purification and identification of novel antioxidant peptides from enzymatic hydrolysates of sardinelle (Sardinellaaurita) by-products proteins

In order to utilise sardinelle (Sardinellaaurita) protein by-products, which is normally discarded as industrial waste in the process of fish manufacturing, heads and viscera proteins were hydrolysed by different proteases to obtain antioxidative peptides. All hydrolysates showed different degrees of hydrolysis and varying degrees of antioxidant activities. Hydrolysate generated with crude enzyme extract from sardine (Sardinapilchardus) displayed high antioxidant activity, and the higher DPPH radical-scavenging activity (87 ± 2.1% at 2 mg/ml) was obtained with a degree of hydrolysis of 6%. This hydrolysate was fractionated by size exclusion chromatography on a Sephadex G-25 into eight major fractions (P₁–P₈). Fraction P₄, which exhibited the highest DPPH scavenging activity, was then fractionated by reversed-phase high performance liquid chromatography (RP-HPLC). Seven antioxidant peptides were isolated. The molecular masses and amino acids sequences of the purified peptides were determined using ESI-MS and ESI-MS/MS, respectively. Their structures were identified as Leu-His-Tyr, Leu-Ala-Arg-Leu, Gly-Gly-Glu, Gly-Ala-His, Gly-Ala-Trp-Ala, Pro-His-Tyr-Leu and Gly-Ala-Leu-Ala-Ala-His. The first peptide displayed the highest DPPH radical-scavenging activity (63 ± 1.57%; at 150 μg/ml) among these peptides.     

Antioxidant, antiacetylcholinesterase and antimicrobial activities of Cymbopogon schoenanthus L. Spreng (lemon grass) from Tunisia

Aqueous extract, proanthocyanidin rich extract, and organic extracts of Cymbopogon schoenanthus L. Spreng (lemon grass) shoots from three different locations in South Tunisia were screened for their antioxidant, acetylcholinesterase and antimicrobial activities. In addition to the evaluation of these activities, the contents of flavonoids and total phenolic compounds were determined.Antioxidant activity measured by DPPH assay showed that the proanthocyanidin extract exhibited higher antioxidant activity than the aqueous extract. Extract concentration providing 50% inhibition (IC₅₀) ranged from 16.4 ± 6.8 μg/mL to 26.4 ± 6.8 μg/mL. The antioxidant activity was also determined using the β-carotene/linoleic acid bleaching test. The best results (IC₅₀ = 0.11 ± 0.10 mg/mL) were obtained with the proanthocyanidin extract of the plants collected from the desert region (Dhibat).The greatest acetylcholinesterase inhibitory activity (IC₅₀ = 0.23 ± 0.04 mg/mL) was exhibited by the ethyl acetate and methanol extracts of the plants collected from the mountainous region. It seems that extracts obtained with more polar solvents gave better results.The proanthocyanidin extracts showed a good antimicrobial activity against Streptococcus sobrinus at low concentration (MIC = 4 mg/mL). Therefore, these extracts could be used to prevent carious lesions by inhibiting S. sobrinus growth.     

Identification and characterization of a novel fermented substance produced by edible Aspergillus oryzae AO-1 that inhibits DPP-IV activity

We identified a novel fermented substance (FKA) produced by the edible Aspergillus oryzae strain AO-1 that inhibited dipeptidyl peptidase IV (DPP-IV) (IC₅₀ = 3.41 mg·mL^− 1). HPLC analysis of FKA showed specific one metabolite (WYK-1), which inhibited DPP-IV (IC₅₀ = 6.98 μM). WYK-1 was identified as a tetrahydroxyisoquinoline derivative. Interestingly, we examined 60 strains of A. oryzae, AO-1 was the only A. oryzae strain that produces WYK-1. This is the first report that an edible A. oryzae strain produces a DPP-IV inhibitor such as WYK-1. This study also suggests that FKA has applications in the development of novel antihyperglycemic therapeutics or functional foods.     

Rapid,simple and sensitive determination of the apparent formation constants of trans-resveratrol complexes with natural cyclodextrins in aqueous medium using HPLC

The study of the complexation of trans-resveratrol with natural cyclodextrins (CDs) in aqueous medium under different physico-chemical conditions of pH or temperature is essential if this antioxidant compound is to be used successfully in the food industry as ingredient of functional foods, due its poor stability, bioavailability and solubility. In this paper, a rapid, simple and sensitive determination of the apparent formation constant of trans-resveratrol/CD complexes by HPLC in aqueous medium has been investigated for first time. It can be observed that trans-resveratrol forms a 1:1 complex with α-, β- and γ-CD. The highest value of the apparent formation constant (K_F = 1922 ± 89 M⁻¹) was found for β-CD and a strong dependence of K_F on pH can be seen in the region where the trans-resveratrol begins the deprotonation of their hydroxyl groups. Moreover, an increase in the system’s temperature produced a decrease in the values of K_F. Finally, to gain information on the mechanism of the trans-resveratrol affinity for CD, the thermodynamic parameters of the complexation were obtained.     

Angiotensin I-converting enzyme-inhibitory peptide fractions from albumin 1 and globulin as obtained of amaranth grain

A number of biopeptides promoting health benefits have been isolated from food-protein hydrolysates and can be released during enzymatic digestion. Antihypertensive peptides can be part of protein fractions from amaranth grain. The objective of this work was to obtain ACE-inhibitory peptide fractions from albumin 1 and the globulin of amaranth (Amaranthus hypochondriacus) grain. Albumin 1 and globulin were hydrolysed with alcalase; hydrolysis was monitored by proteolytic degradation and by ACE-inhibitory activity. The highest ACE-inhibitory activity was 40% and 35% as obtained after 18 and 15 h hydrolysis for albumin 1 and globulin, respectively. Further separation and purification of the ACE-inhibitory peptide fractions were carried out by gel filtration and C₁₈ RP–HPLC. The IC₅₀ was 0.35 ± 0.02 mg/ml for albumin 1 peptide fraction and 0.15 ± 0.03 mg/ml for globulin peptide fraction. Albumin 1 peptide fraction showed an competitive mode of ACE inhibition, whereas the globulin peptide fraction was competitive. The globulin peptide fraction may have one of the most active naturally-occurring ACE-inhibitory peptides.     

Determination of stilbenes in Sicilian pistachio by high-performance liquid chromatographic diode array (HPLC-DAD/FLD) and evaluation of eventually mycotoxin contamination

In this work, we have investigated about the presence of several natural stilbenes in 12 samples of pistachios harvested from 10 different farms of Sicily (Bronte and Agrigento). At the same time, we have evaluated the relation between the stilbenes synthesis and the possible contamination of mycotoxin produced by Aspergillus flavus and Aspergillus parasiticus. We have found two types of stilbenes in the samples of pistachios examined: trans-resveratrol and trans-resveratrol-3-O-β-glucoside (trans-piceid). Their concentration ranged from 0.07 to 0.18 mg/kg (av. = 0.12 ± 0.03 mg/kg) for trans-resveratrol, from 6.20 to 8.15 mg/kg (av. = 6.97 ± 0.55 mg/kg) for trans-piceid and from 6.38 to 8.27 mg/kg (av. = 7.09 ± 0.54 mg/kg) for total resveratrol.     

Use of high-concentration-short-time chlorine dioxide gas treatments for the inactivation of Salmonella enterica spp. inoculated onto Roma tomatoes

Salmonella outbreaks have been recently linked to the consumption of fresh tomatoes. Thus, there is a need to develop systems that reduce the risk of microbial contamination to increase product shelf-life and keep fresh fruit attributes. The objectives of this study were to evaluate high-concentration-short-time chlorine dioxide gas treatments effects on Salmonella-inoculated Roma tomatoes and determine the optimal treatment conditions for microbial inactivation and shelf-life extension. Effects of ClO₂ concentration (2, 5, 8 and 10 mg/l) and exposure time (10, 30, 60, 120 and 180 s) on inoculated Roma tomatoes were studied. Salmonella enterica strains, serotype Montevideo, Javiana and Baildon, were used to experimentally inoculate the food product. After ClO₂ treatments, tomatoes were stored at room temperature for 28 days. Inherent microbial population, change in tomato color, and chlorine dioxide gas residuals were evaluated. ANOVA analysis showed that both ClO₂ concentration and treatment time were significant (p < 0.01) for Salmonella inactivation. Surviving Salmonella populations of 3.09, 2.17 and 1.16 log CFU/cm² were obtained treating tomatoes with 8 mg/l ClO₂ for 60 s, 10 mg/l ClO₂ for 120 s, and 10 mg/l for 180 s, respectively (initial Salmonella population: 6.03 ± 0.11 log CFU/cm²). The selected treatments significantly reduced background microflora (p < 0.05), while fruit color and residual contents were not significantly different (p > 0.05), as compared to the control. Results suggest the potential for high-concentration-short-time treatments ClO₂ gas as an effective pathogen inactivation technology for large-scale produce packing operations.     

Value-added utilization of yak milk casein for the production of angiotensin-I-converting enzyme inhibitory peptides

Yak (Bos grunniens) milk casein derived from Qula, a kind of acid curd cheese from northwestern China, was hydrolysed with alcalase. The hydrolysates collected at different hydrolysis times (0 min, 60 min, 120 min, 180 min, 240 min, 300 min, 360 min) were assayed for the inhibitory activity of angiotensin-I-converting enzyme (ACE), and the one obtained at 240 min hydrolysis showed the highest ACE inhibitory activity. The active hydrolysate was further consecutively separated by ultrafiltration with 10 kDa and then with 6 kDa molecular weight cut-off membranes into different parts, and the 6 kDa permeate showed the highest ACE-inhibiting activity. This active fraction was further purified to yield two novel ACE-inhibiting peptides, whose amino acid sequences were Pro–Pro–Glu–Ile–Asn (PPEIN)(κ-CN; f156–160) and Pro–Leu–Pro–Leu–Leu (PLPLL) (β-CN; f136–140), respectively. The molecular weight and IC₅₀ value of the peptides were 550 Da and 566.4 Da, and 0.29 ± 0.01 mg/ml and 0.25 ± 0.01 mg/ml, respectively.     

Inhibitory effects of p-alkylbenzoic acids on the activity of polyphenol oxidase from potato (Solanum tuberosum)

In order to prevent potato from browning during preservation and processing, the effects of p-alkylbenzoic acids (p-propylbenzoic acid, p-butylbenzoic acid, p-pentylbenzoic acid, p-hexylbenzoic acid, p-heptylbenzoic acid, p-octylbenzoic acid) on the activity of potato polyphenol oxidase (PPO) have been studied. The PPO was extracted with PBS (pH 6.8), fractionated with ammonium sulphate, concentrated, and purified with Sephadex G-100 (1.8 × 100 cm) filtration chromatography. The active fractions were pooled and the PPO activity was determined to be 79.83 U/mg. The results showed that p-alkylbenzoic acids could strongly inhibit the activity of potato PPO. The values of IC₅₀ of these six inhibitors were determined to be 0.213, 0.180, 0.152, 0.106, 0.075 and 0.047 mM, respectively. Kinetic analyses showed that p-alkylbenzoic acids used in this experiment were reversible and noncompetitive inhibitors to the enzyme. The inhibitory effects were potentiated with increasing lengths of the hydrocarbon chains, indicating that the inhibitory efficiency on the enzyme was influenced by the steric effect of the substituted groups.     

Quality assessment of wild European eel (Anguilla anguilla) stored in ice

The quality assessment of wild European eel (Anguilla anguilla) stored in ice and in boxes without ice (3 ± 1 °C) was investigated by the sensory analysis, levels of nucleotide breakdown products and biogenic amines for up to 19 days. Sensory analysis was assessed using the Tasmanian Food Research Unit Scheme. K and related values (K_i, G, P, H and F_r) were used as freshness indicators. Linear regressions (r²) obtained from K, K_i, G, P, H and F_r were 0.95, 0.96, 0.83, 0.96, 0.99 and 0.96, respectively, for eel stored in ice whereas, for eel kept in boxes without ice, the values were 0.86, 0.86, 0.96, 0.91, 0.98 and 0.86, respectively. When eel stored in ice and in boxes without ice were considered at the limit of acceptability by assessors at ∼12–14 days and ∼5–7 days, respectively, the average K, K_i and P values were ∼70–85%, H values were ∼60% and F_r values were ∼10% for both storage conditions. The level of histamine exceeded the legal limit (5 mg/100 g fish) in eel stored without ice after 6–7 days and, in ice, after 13–14 days of storage, at which time eels were rejected by the sensory panel. The concentrations of biogenic amines were higher in eel stored in boxes without ice than in eel kept in ice. The levels of histamine in the muscle of eel kept in boxes without ice and in ice increased to the maximum levels of 17.9 mg/100 g on day 12 and 12.6 mg/100 g on day 19, respectively.